Photo-oxidation of cardiolipin and cytochrome c with bilayer-embedded Pc 4.

Singlet oxygen, (1)O(2), is produced by absorption of red light by the phthalocyanine dye Pc 4, followed by energy transfer to dissolved triplet molecular oxygen, (3)O(2). In tissues, Pc 4 concentrates in lipid bilayers, and particularly in mitochondrial membranes, because of its positive charge. Illumination of cells and tissues with red light after uptake of Pc 4 results in cell death. The potential initial chemical steps that result in cellular dysfunction have been characterized in this study. Both unsaturated acyl chains of phospholipids and proteins are identified as targets of oxidation. Tetra-linoleoyl cardiolipin was oxidized in both liposomes and mitochondria after Pc 4-mediated (1)O(2) generation. Evidence for the formation of both mono- and bis-hydroperoxide adducts of single linoleoyl side chains is provided by ESI-MS and ESI-MS/MS. Similarly, illumination of Pc 4 in liposomes and mitochondria resulted in cytochrome c oxidation as detected by oxidation of His 26 in the peptide H(26)*KTGPNLHGLFGK, further supporting the potential use of this peptide as a biomarker for the presence of mitochondrial oxidative stress characteristic of (1)O(2) in vivo (J. Kim et al., Free Radic. Biol. Med. 44:1700-1711; 2008). These observations provide evidence that formation of lipid hydroperoxides and/or protein oxidation can be the initial chemical steps in Pc 4-mediated induction of apoptosis in photodynamic therapy.

Radiation-induced fragmentation of cardiolipin in a model membrane.

PURPOSE: To obtain evidence for the possibility of free-radical fragmentation of cardiolipin under the action of ionizing radiation as measured by its aqueous dispersion from liposomes. MATERIALS AND METHODS: Liposomes of tetramyristoylcardiolipin (TMCL) were exposed to gamma-rays from 60Co or 137Cs sources at doses between 1 and 24kGy. Fragmentation products were identified using thin-layer chromatography and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). RESULTS: Using MALDI-TOF MS and thin-layer chromatography, it was shown that gamma-irradiation of liposomes consisting of TMCL was accompanied by free-radical fragmentation of the lipid to form dimiristoylphosphatidic acid and dimiristoylphosphatidyl hydroxyacetone. The yields of dimiristoylphosphatidic acid were greater than those of dimiristoylphosphatidyl hydroxyacetone, and formation of the named compounds was inhibited by dissolved oxygen. CONCLUSION: It is shown for the first time that on gamma-irradiation, cardiolipin can undergo free-radical fragmentation in its polar component.

Role of endogenous lipids in the chromophore regeneration of bacteriorhodopsin.

The regeneration method of Khorana [J. Biol. Chem. 262 (1987) 9271] has been modified in order to study the effect of endogenous archaeabacterial lipids and, in particular, of glycocardiolipin (GlyC) in the refolding and chromophore regeneration of bacteriorhodopsin (BR). BR refolding and chromophore regeneration could be obtained in the presence of endogenous lipid mixtures containing or not containing glycocardiolipin; however, the kinetics of bacteriorhodopsin regeneration in the presence of glycocardiolipin was faster than in its absence. These results show for the first time that the interaction of glycocardiolipin with bacteriorhodopsin favours its refolding from the denaturated state and the chromophore regeneration.

[Opposite effect of lipofuscin granules and melanosomes from human retinal pigment epithelium of eye on photooxidation of cardiolipin]. / Raznonapravlennost' deistviia lipofustsinovykh granul i melanosom iz retinal'nogo pigmentnogo épiteliia glaza cheloveka pri fotookislenii kardiolipina.

The influence of lipofuscin granules and melanosomes from human retinal pigment epithelium on the light-induced photooxidation of cardiolipin liposomes and the generation of superoxide radicals was studied. Lipofuscin granules were able to stimulate, while melanosomes inhibited, the cardiolipin photooxidation. The visible light irradiation of both melanosomes and lipofuscin granules generated superoxide radicals with mean rates of 1.5 nmole/min/10(7) and 38 nmole/min/10(7) granules, accordingly. However, melanosomes but not lipofuscin granules reacted readily with superoxide radicals. Moreover, the rate constant of degradation of superoxide radicals in the presence of melanosomes was about five orders of magnitude higher than the rate constant of its photogeneration. Therefore, we propose that melanosomes in retinal pigment epithelium cells have a photoprotective role whereas lipofuscin granules may stimulate photodestructive reactions.

Antioxidative activity of synthetic melanins. Cardiolipin liposome model.

The inhibiting effect of melanin synthesized from dihydroxyphenylalanine (DOPA), dopamine, adrenaline and adrenolutin on the ultraviolet- or the Fe(2+)-ascorbic acid-induced peroxidation of cardiolipin liposomes has been studied. All these melanins are able to inhibit both the ultraviolet- and the Fe(2+)-ascorbic acid-induced lipid peroxidation. Antioxidative activity of melanins enhances in the order: dopamine-melanin less than melanin synthesized from dopamine in the presence of Cu(2+) less than DOPA--melanin less than melanin synthesized from adrenaline in the presence of Cu(2+) approximately equal to adrenolutin-melanin, and correlates with their ability to scavenge superoxide anion radical. The optical screening effect of the investigated melanins in the inhibition of lipid peroxidation was not higher than 15% for the most active melanins.

[Inhibition of UV-induced accumulation of lipid peroxides by melanins and ommochromes]. / Ingibirovanie UF-indutsirovannoi akkumuliatsii lipidnykh gidroperoksidov melaninami i ommokhromami.

The effect of screening pigments--melanin and ommochromes--on the UV-induced accumulation of lipid hydroperoxides by cardiolipin was studied. It was shown that the time dependence of lipid hydroperoxide accumulation at different UV irradiation intensities is linear. Addition of melanin or ommochromes results in a significant decrease of the lipid hydroperoxide accumulation rate. The kinetic curve rapidly reaches the saturation values, which is suggestive of intensive chemical inhibition of the reaction. The screens transmitting UV irradiation in the same degree as do melanin and ommochromes have a far less pronounced inhibiting effect. The inhibitory mechanism of screening pigments based on their antiradical activity is discussed.

[The system of protection of the eye structures from photo damage. Screening pigments in vertebrates--melanosomes as inhibitors of photo-oxidation processes]. / Sistema zashchity struktur glaza ot fotopovrezhdeniia. Ekraniruiushchie pigmenty pozvonochnykh--melanosomy kak ingibitory protsessov fotookisleniia.

Melanosomes from retinal pigmented epithelium readily inhibit photooxidation of lipids. This effect is due to both passive screening of light and chemical interaction of melanosomes with products of oxidation. The role of melanosomes is discussed in the system of optic and chemical protection of eye structures from photo injury. It is suggested that in other tissues (skin) melanosomes account for the similar function of optic and chemical (antioxidative) protection from the injurious effect of UV and visible light.

[Effect of changes in the lipids of mitochondrial membranes on the activity of Mg2+-dependent ATPase]. / Vliianie izmenenii v lipidakh membran mitokhondrii na aktivnost' Mg2+-zavisimoi ATFazy.

The results obtained permit to assume that irradiation causes dysfunction of the regulatory system that provides the interdependence between the antioxidative activity of lipids, their composition and the activity of membrane-bound enzymes. There is virtually no correlation between the changes in hydrolase activity of the enzyme and the quantity of phosphatidylethanolamine (PE) and cardiolipin (CL). During the first hours following irradiation the dependence between the changes in the synthetase activity of ATPase and the fluidity of the lipid component of the membrane is directly proportional (just as it is observed in normal conditions); the lesser the fluidity of the lipid component the higher the hydrolase activity of the enzyme.